The unique environment of the membrane protein has caused a severe discrepancy between the level of importance of membrane proteins and the level of our current understanding of them. To rectify this disparity we have developed a novel method, The Steric Trap Method, which couples protein unfolding to a measurable binding event. The simultaneous binding of two monovalent streptavidin (mSA) molecules to a dually, biotinylated target protein is inhibited when the protein is folded due to steric hindrance created by the bulky bound molecules and can occur only when an unfolded state is sampled under the condition of dynamic equilibrium. Therefore, the binding affinity of mSA for the second biotin site on the target protein is correlated to the unfolding energy of the protein. This allows the investigation of folding energetics in the native environment of lipid bilayers without the harsh denaturing conditions utilized by existing methods. It also eliminates the need for dilution, a disadvantage of existing methods, essential for probing high affinity interactions.

We have successfully applied the steric trap method to study both soluble protein folding for the model enzyme, dihydrofolate reducatase (DHFR), and membrane protein folding for glycophorin A (GpA) and diacylglycerol kinase (DGK). We have used the steric trap method to study protein folding and stability, protein-protein interaction energetics, and unfolding kinetics. We can investigate the effects of different mutations, detergents, lipids, or any other change to the folding conditions. Lastly, we can also irreversibly (on the order of several days) trap the unfolded state to enable its characterization.